The overall aims are to gain a better understanding of the mechanisms of differential targeting of neurons by prion strains, a better understanding of the cellular and molecular mechanisms of nerve cell dysfunction, degeneration and death in prion diseases, and to test whether clearance of PrpSc from the brain corrects cellular defects and neuronal vacuolation and foster reactive synaptogenesis. Aim 1 tests a membrane hypothesis of pathogenesis that proposes theat the most fundamental cellular defects in prion diseases are an accumulation of abnormal PrP (PrP/Sc in scrapie and CJD and tm PrP in some GSS-like disease) in neuronal plasma membranes with a proportional change in the properties of the plasma membrane. We will correlate the degree of change in synaptosomal membrane fluidity, measured by electron spin resonance spectroscopy, with the concentration of PrPSc or tmPrP. In Aim 2 we will test the hypothesis proposed by others that dysfunction and possibly death of the parvalbumin subset of GABAergic neurons precedes grey matter vacuolation. An extensive stereological quantitative study of parvalbumin and GABA nerves cell bodies and synapse numbers as a function of PrPSc or tmPrP accumulation is proposed. Aim 3 proposes to use doxycycline-treated Tg(tTA:MoPrP) mice (Project 3) to directly test whether PrPSc accumulation in plasma membrane is the cause of neuronal dysfunction, specifically by testing the effect of clearance of PrPSc on plasma membrane properties, vacuolation, and parvalbumin neurons. We will also test whether clearance of PrPSc corrects decreased release of GABA from synaptosomes (see Progress Report) and allows reactive synaptogenesis to replace lost synapses. Aim 4 focuses on our hypothesis that the two Asn-linked PrP oligosaccharides influence host animal and prion-strain determined differential targeting of neurons. Specifically, we will examine a new transgenic mouse the effect on the disease phenotype. We will also test whether or not prion strain properties are influenced by the brain region from which they are derived by transmitting human vCJD derived from different brain regions to susceptible transgenic mice.